Fatty liver disease (FLD) is a burgeoning health problem that affects one-third of adults and an increasing number of children in the U.S. The two most common forms of FLD are nonalcoholic liver disease (NAFLD) and alcoholic liver disease (ALD). Both disorders begin with accumulation of triglyceride (TG) in the liver (steatosis), which in some individuals elicits an inflammatory response [nonalcoholic steatohepatitis (NASH) or alcoholic steatosis (ASH)] that can progress to cirrhosis and liver cancer. Therapeutic options to arrest disease progression in both disorders are very limited. The development of treatments for FLD has been hampered by limited appreciation of the molecular underpinnings of the disease, the lack of noninvasive markers of disease progression in humans, and a paucity of animal models that accurately recapitulate the pathogenesis of human FLD. As a first step to address these obstacles, our group used human genetics to identify genes that contribute to FLD. We identified a missense mutation (I148M) in patatin-like phospholipase domain-containing protein, PNPLA3, which is strongly associated with both hepatic TG content and inflammation. We showed that the variant is very frequent in Hispanics, who have the highest prevalence of NAFLD in the United States. Over 50 independent studies have confirmed and extended our findings to show that PNPLA3-I148M is enriched in individuals with biopsy-proven steatohepatitis, cirrhosis, and hepatocellular carcinoma both due to ALD and NAFLD. Thus, PNPLA3 is implicated as a contributing factor in the full spectrum of both NAFLD and ALD, suggesting that these two forms of FLD share not only pathological features but also molecular mechanisms in common. Recently, we identified a genetic variant in a second gene, TM6SF2, that confers susceptibility to both steatosis and inflammation. A missense variant in this gene is associated with increased hepatic TG content independently of PNPLA3. The function of this gene is unknown. In this application we will use a combination of classical biochemistry and physiology plus state-of-the-art mass spectrometry in mice with genetically-defined changes in PNPLA3 and TM6SF2 function to elucidate the physiological roles of these two proteins and the molecular mechanisms by which they promote hepatic fat accumulation and inflammation. We will leverage these findings to develop mouse models that more accurately recapitulate human NAFLD and ALD and provide improved reagents for pre-clinical testing. RELEVANCE: By elucidating the biological roles of PNPLA3 and TM6SF2 and the mechanisms by which variants in these proteins confer susceptibility to FLD, the experiments outlined in this proposal will provide new insights into the pathogenesis of a major human disease that continues to increase in prevalence. Our ultimate goal is to develop new approaches and strategies to diagnose, prevent, and treat FLD.